The proposed research will be a systematic biochemical investigation of the mechanism and regulation of Escherichia coli RNA polymerase. The interaction of this enzyme with its DNA template will be probed by functional and spectroscopic experiments in vitro. The focus throughout will be to understand how DNA sequence, protein activators, and small molecules affect the rate limiting steps in RNA chain initiation. The biochemical basis of promoter mutations will be thoroughly studied. The mechanism of transcription activation will be investigated on the lac operon. We also want to know how ribosomal RNA and tRNA synthesis is controlled in E. coli. Antitermination of transcription as a regulatory process will be investigated by determining the mechanism of the bacteriophage lambda N protein. The answers to the questions we have posed will contribute to the biochemical description of the control of gene expression.